Role of gelatinases MMP-2 and MMP-9 in tissue remodeling following acute lung injury

Acute lung injury is characterized by a severe disruption of alveolo-capillary structures and includes a variety of changes in lung cell populations. Evidence suggests the occurrence of rupture of the basement membranes and interstitial matrix remodeling during acute lung injury. The dynamic equilibrium of the extracellular matrix (ECM) under physiological conditions is a consequence of the balance between the regulation of synthesis and degradation of ECM components. Matrix metalloproteinases (MMPs) represent a group of enzymes involved in the degradation of most of the components of the ECM and therefore participate in tissue remodeling associated with pathological situations such as acute lung injury. MMP activity is regulated by proteolytic activation of the latent secreted proenzyme and by interaction with specific tissue inhibitors of metalloproteinases. This review details our knowledge of the involvement of MMPs, namely MMP-2 and MMP-9, in acute lung injury and acute respiratory distress syndrome.

Cycle modulation of insulin-like growth factor-binding protein 1 in human endometrium

Endometrium is one of the fastest growing human tissues. Sex hormones, estrogen and progesterone, in interaction with several growth factors, control its growth and differentiation. Insulin-like growth factor 1 (IGF-1) interacts with cell surface receptors and also with specific soluble binding proteins. IGF-binding proteins (IGF-BP) have been shown to modulate IGF-1 action. Of six known isoforms, IGF-BP-1 has been characterized as a marker produced by endometrial stromal cells in the late secretory phase and in the decidua. In the current study, IGF-1-BP concentration and affinity in the proliferative and secretory phase of the menstrual cycle were measured. Endometrial samples were from patients of reproductive age with regular menstrual cycles and taking no steroid hormones. Cytosolic fractions were prepared and binding of 125I-labeled IGF-1 performed. Cross-linking reaction products were analyzed by SDS-polyacrylamide gel electrophoresis (7.5%) followed by autoradiography. 125I-IGF-1 affinity to cytosolic proteins was not statistically different between the proliferative and secretory endometrium. An approximately 35-kDa binding protein was identified when 125I-IGF-1 was cross-linked to cytosol proteins. Secretory endometrium had significantly more IGF-1-BP when compared to proliferative endometrium. The specificity of the cross-linking process was evaluated by the addition of 100 nM unlabeled IGF-1 or insulin. Unlabeled IGF-1 totally abolished the radioactivity from the band, indicating specific binding. Insulin had no apparent effect on the intensity of the labeled band. These results suggest that IGF-BP could modulate the action of IGF-1 throughout the menstrual cycle. It would be interesting to study this binding protein in other pathologic conditions of the endometrium such as adenocarcinomas and hyperplasia.

The brain decade in debate: VIII. Peptide hormones and behavior: cholecystokinin and prolactin

This article is a transcription of an electronic symposium held on November 28, 2000 in which active researchers were invited by the Brazilian Society of Neuroscience and Behavior (SBNeC) to discuss the advances of the last decade in the peptide field with particular focus on central actions of prolactin and cholecystokinin. The comments in this symposium reflect the diversity of prolactin and cholecystokinin research and demonstrate how the field has matured. Since both peptides play a role in reproductive behaviors, particularly mother-infant interactions, this was the starting point of the discussion. Recent findings on the role of the receptor subtypes as well as interaction with other peptides in this context were also discussed. Another issue discussed was the possible role of these peptides in dopamine-mediated rewarding systems. Both prolactin and cholecystokinin are involved in mechanisms controlling food intake and somatic pain thresholds. The role of peripheral inputs through vagal afferents modulating behavior was stressed. The advent of knockout animals as potential generators of new knowledge in this field was also addressed. Finally, interactions with other neuropeptides and investigation of the role of these peptides in other fields such as immunology were mentioned. Knowledge about the central functions of prolactin and cholecystokinin has shown important advances. The role of these peptides in neurological and psychiatric syndromes such as anorexia, drug abuse and physiological disturbances that lead to a compromised maternal behavior seems relevant.

Evaluation of the cholinomimetic actions of trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana (Gastropoda, Opisthobranchia)

Trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana, negatively modulates vagal response, indicating a probable ability to inhibit cholinergic responses. In the present study, the pharmacological profile of trimethylsulfonium was characterized on muscarinic and nicotinic acetylcholine receptors. In rat jejunum the contractile response induced by trimethylsulfonium (pD2 = 2.46 ± 0.12 and maximal response = 2.14 ± 0.32 g) was not antagonized competitively by atropine. The maximal response (Emax) to trimethylsulfonium was diminished in the presence of increasing doses of atropine (P<0.05), suggesting that trimethylsulfonium-induced contraction was not related to muscarinic stimulation, but might be caused by acetylcholine release due to presynaptic stimulation. Trimethylsulfonium displaced [³H]-quinuclidinyl benzilate from rat cortex membranes with a low affinity (Ki = 0.5 mM). Furthermore, it caused contraction of frog rectus abdominis muscles (pD2 = 2.70 ± 0.06 and Emax = 4.16 ± 0.9 g), which was competitively antagonized by d-tubocurarine (1, 3 or 10 µM) with a pA2 of 5.79, suggesting a positive interaction with nicotinic receptors. In fact, trimethylsulfonium displaced [³H]-nicotine from rat diaphragm muscle membranes with a Ki of 27.1 µM. These results suggest that trimethylsulfonium acts as an agonist on nicotinic receptors, and thus contracts frog skeletal rectus abdominis muscle and rat jejunum smooth muscle via stimulation of postjunctional and neuronal prejunctional nicotinic cholinoreceptors, respectively.

APOA1/C3/A4 gene cluster variability and lipid levels in Brazilian children

Genetic studies have suggested that polymorphisms of genes coding for apolipoproteins are significant determinants of serum lipoprotein and lipid levels in adults. However, only a few studies have investigated the association of these polymorphisms in children. Therefore, in the present investigation we studied the distribution of APOA1 -75 G>A, +83 C>T, APOC3 -482 C>T, -455 T>C and 3238 C>G, and APOA4 Q360H and T347S polymorphisms and their influence on plasma lipoprotein levels in children from a Brazilian northeastern admixed population. The seven polymorphic sites were genotyped in 414 children aged 5 to 15 years (mean 8.9 ± 2.9). The genotypes of the seven polymorphic sites were assessed by PCR-RFLP methods. The frequencies of the less common alleles were, in general, intermediate among parental populations, as expected. Strong linkage disequilibrium was detected between polymorphisms at the APOA1, APOC3 and APOA4 loci in this admixed population sample. Overall the genotype effects seen in adults were weaker or absent in children. The APOC3/-455 and APOA4 T347S variants showed significant effects on HDL cholesterol in girls (P = 0.033 and P = 0.016, respectively). Significantly higher plasma total (P = 0.003) and LDL cholesterol (P = 0.004) levels were observed in boys who were carriers of the 3238G allele at the APOC3/3238 C>G site. These results disclosed an overall absence of associations between these polymorphisms and lipids in children. This finding is not unexpected because expression of the effect of these polymorphisms might depend on the interaction with environmental variables both internal and external to the individual.

Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

Enveloped viruses always gain entry into the cytoplasm by fusion of their lipid envelope with a cell membrane. Some enveloped viruses fuse directly with the host cell plasma membrane after virus binding to the cell receptor. Other enveloped viruses enter the cells by the endocytic pathway, and fusion depends on the acidification of the endosomal compartment. In both cases, virus-induced membrane fusion is triggered by conformational changes in viral envelope glycoproteins. Two different classes of viral fusion proteins have been described on the basis of their molecular architecture. Several structural data permitted the elucidation of the mechanisms of membrane fusion mediated by class I and class II fusion proteins. In this article, we review a number of results obtained by our laboratory and by others that suggest that the mechanisms involved in rhabdovirus fusion are different from those used by the two well-studied classes of viral glycoproteins. We focus our discussion on the electrostatic nature of virus binding and interaction with membranes, especially through phosphatidylserine, and on the reversibility of the conformational changes of the rhabdovirus glycoprotein involved in fusion. Taken together, these data suggest the existence of a third class of fusion proteins and support the idea that new insights should emerge from studies of membrane fusion mediated by the G protein of rhabdoviruses. In particular, the elucidation of the three-dimensional structure of the G protein or even of the fusion peptide at different pH's might provide valuable information for understanding the fusion mechanism of this new class of fusion proteins.

Specific structural features of syndecans and heparan sulfate chains are needed for cell signaling

The syndecans, heparan sulfate proteoglycans, are abundant molecules associated with the cell surface and extracellular matrix and consist of a protein core to which heparan sulfate chains are covalently attached. Each of the syndecan core proteins has a short cytoplasmic domain that binds cytosolic regulatory factors. The syndecans also contain highly conserved transmembrane domains and extracellular domains for which important activities are becoming known. These protein domains locate the syndecan on cell surface sites during development and tumor formation where they interact with other receptors to regulate signaling and cytoskeletal organization. The functions of cell surface heparan sulfate proteoglycan have been centered on the role of heparan sulfate chains, located on the outer side of the cell surface, in the binding of a wide array of ligands, including extracellular matrix proteins and soluble growth factors. More recently, the core proteins of the syndecan family transmembrane proteoglycans have also been shown to be involved in cell signaling through interaction with integrins and tyrosine kinase receptors.

The extracellular matrix provides directional cues for neuronal migration during cerebellar development

Normal central nervous system development relies on accurate intrinsic cellular programs as well as on extrinsic informative cues provided by extracellular molecules. Migration of neuronal progenitors from defined proliferative zones to their final location is a key event during embryonic and postnatal development. Extracellular matrix components play important roles in these processes, and interactions between neurons and extracellular matrix are fundamental for the normal development of the central nervous system. Guidance cues are provided by extracellular factors that orient neuronal migration. During cerebellar development, the extracellular matrix molecules laminin and fibronectin give support to neuronal precursor migration, while other molecules such as reelin, tenascin, and netrin orient their migration. Reelin and tenascin are extracellular matrix components that attract or repel neuronal precursors and axons during development through interaction with membrane receptors, and netrin associates with laminin and heparan sulfate proteoglycans, and binds to the extracellular matrix receptor integrins present on the neuronal surface. Altogether, the dynamic changes in the composition and distribution of extracellular matrix components provide external cues that direct neurons leaving their birthplaces to reach their correct final location. Understanding the molecular mechanisms that orient neurons to reach precisely their final location during development is fundamental to understand how neuronal misplacement leads to neurological diseases and eventually to find ways to treat them.

Transient outward potassium current and Ca2+ homeostasis in the heart: beyond the action potential

Normal central nervous system development relies on accurate intrinsic cellular programs as well as on extrinsic informative cues provided by extracellular molecules. Migration of neuronal progenitors from defined proliferative zones to their final location is a key event during embryonic and postnatal development. Extracellular matrix components play important roles in these processes, and interactions between neurons and extracellular matrix are fundamental for the normal development of the central nervous system. Guidance cues are provided by extracellular factors that orient neuronal migration. During cerebellar development, the extracellular matrix molecules laminin and fibronectin give support to neuronal precursor migration, while other molecules such as reelin, tenascin, and netrin orient their migration. Reelin and tenascin are extracellular matrix components that attract or repel neuronal precursors and axons during development through interaction with membrane receptors, and netrin associates with laminin and heparan sulfate proteoglycans, and binds to the extracellular matrix receptor integrins present on the neuronal surface. Altogether, the dynamic changes in the composition and distribution of extracellular matrix components provide external cues that direct neurons leaving their birthplaces to reach their correct final location. Understanding the molecular mechanisms that orient neurons to reach precisely their final location during development is fundamental to understand how neuronal misplacement leads to neurological diseases and eventually to find ways to treat them.

The role of autolysis loop in determining the specificity of coagulation proteases

We recently demonstrated that the substitution of the autolysis loop (residues 143 to 154 in the chymotrypsin numbering system) of activated protein C (APC) with the corresponding loop of factor Xa (fXa) renders the APC mutant (APC/fX143-154) susceptible to inhibition by antithrombin (AT) in the presence of pentasaccharide. Our recent results further indicated, that in addition to an improvement in the reactivity of APC/fX143-154 with AT, both the amidolytic and anti-factor Va activities of the mutant APC have also been significantly increased. Since the autolysis loop of APC is five residues longer than the autolysis loop of fXa, it could not be ascertained whether this loop in the mutant APC specifically interacts with the activated conformation of AT or if a shorter autolysis loop is responsible for a global improvement in the catalytic activity of the mutant protease. To answer this question, we prepared another APC mutant in which the autolysis loop of the protease was replaced with the corresponding loop of trypsin (APC/Tryp143-154). Unlike an ~500-fold improvement in the reactivity of APC/fX143-154 with AT in the presence of pentasaccharide, the reactivity of APC/Tryp143-154 with the serpin was improved ~10-fold. These results suggest that both the length and structure of residues of the autolysis loop are critical for the specificity of the coagulation protease interaction with AT. Further factor Va inactivation studies with the APC mutants revealed a similar role for the autolysis loop of APC in the interaction with its natural substrate.

Pharmacology and toxicology of diphenyl diselenide in several biological models

The pharmacology of synthetic organoselenium compounds indicates that they can be used as antioxidants, enzyme inhibitors, neuroprotectors, anti-tumor and anti-infectious agents, and immunomodulators. In this review, we focus on the effects of diphenyl diselenide (DPDS) in various biological model organisms. DPDS possesses antioxidant activity, confirmed in several in vitro and in vivo systems, and thus has a protective effect against hepatic, renal and gastric injuries, in addition to its neuroprotective activity. The activity of the compound on the central nervous system has been studied since DPDS has lipophilic characteristics, increasing adenylyl cyclase activity and inhibiting glutamate and MK-801 binding to rat synaptic membranes. Systemic administration facilitates the formation of long-term object recognition memory in mice and has a protective effect against brain ischemia and on reserpine-induced orofacial dyskinesia in rats. On the other hand, DPDS may be toxic, mainly because of its interaction with thiol groups. In the yeast Saccharomyces cerevisiae, the molecule acts as a pro-oxidant by depleting free glutathione. Administration to mice during cadmium intoxication has the opposite effect, reducing oxidative stress in various tissues. DPDS is a potent inhibitor of d-aminolevulinate dehydratase and chronic exposure to high doses of this compound has central effects on mouse brain, as well as liver and renal toxicity. Genotoxicity of this compound has been assessed in bacteria, haploid and diploid yeast and in a tumor cell line.

Effect of oral ingestion of an extract of the herb Uncaria tomentosa on the biodistribution of sodium pertechnetate in rats

The aim of the present study was to determine the effect of the oral ingestion of an extract of the herb Uncaria tomentosa (cat's claw) on the biodistribution of the radiobiocomplex sodium pertechnetate (Na99mTcO4) in rats. The animals (male Wistar rats, 2 months old, 180-220 g), were treated (1 mL) with an U. tomentosa extract (32 mg/mL, N = 5) or 0.9% NaCl solution (control, N = 5) for 7 days. After this period, Na99mTcO4 (3.7 MBq, 0.3 mL) was injected through the ocular plexus and after 10 min the rats were killed, the organs isolated and counted in a well-gamma counter. A significant (P < 0.05) alteration in Na99mTcO4 uptake i) from 0.57 ± 0.008 to 0.39 ± 0.06 %ATI/organ (P < 0.05) and from 0.57 ± 0.17 to 0.39 ± 0.14 %ATI/g (P < 0.05) was observed in the heart, ii) from 0.07 ± 0.02 to 0.19 ± 0.07 %ATI/g in the pancreas, and iii) from 0.07 ± 0.01 to 0.18 ± 0.07 %ATI/g (P < 0.05) in muscle after treatment with this extract. Although these results were obtained with animals, caution is advisable in the interpretation of the nuclear medicine examination when the patient is using this herb. This finding is probably an example of drug interaction with a radiopharmaceutical, a fact that could lead to misdiagnosis of the examination in clinical practice with unexpected consequences for the patient.

Noncrystalline uric acid inhibits proteoglycan and glycosaminoglycan synthesis in distal tubular epithelial cells (MDCK)

Hyperuricemia is associated with renal stones, not only consisting of uric acid (UrAc) but also of calcium oxalate (CaOx). Glycosaminoglycans (GAGs) are well-known inhibitors of growth and aggregation of CaOx crystals. We analyzed the effect of noncrystalline UrAc on GAG synthesis in tubular distal cells. MDCK (Madin-Darby canine kidney) cells were exposed to noncrystalline UrAc (80 µg/mL) for 24 h. GAGs were labeled metabolically and characterized by agarose gel electrophoresis. The expression of proteoglycans and cyclooxygenase 2 (COX-2) was assessed by real-time PCR. Necrosis, apoptosis and prostaglandin E2 (PGE2) were determined by acridine orange, HOESCHT 33346, and ELISA, respectively. CaOx crystal endocytosis was evaluated by flow cytometry. Noncrystalline UrAc significantly decreased the synthesis and secretion of heparan sulfate into the culture medium (UrAc: 2127 ± 377; control: 4447 ± 730 cpm) and decreased the expression of perlecan core protein (UrAc: 0.61 ± 0.13; control: 1.07 ± 0.16 arbitrary units), but not versican. Noncrystalline UrAc did not induce necrosis or apoptosis, but significantly increased COX-2 and PGE2 production. The effects of noncrystalline UrAc on GAG synthesis could not be attributed to inflammatory actions because lipopolysaccharide, as the positive control, did not have the same effect. CaOx was significantly endocytosed by MDCK cells, but this endocytosis was inhibited by exposure to noncrystalline UrAc (control: 674.6 ± 4.6, CaOx: 724.2 ± 4.2, and UrAc + CaOx: 688.6 ± 5.4 geometric mean), perhaps allowing interaction with CaOx crystals. Our results indicate that UrAc decreases GAG synthesis in MDCK cells and this effect could be related to the formation of UrAc and CaOx stones.

Effect of addition of different hydrocolloids on pasting, thermal, and rheological properties of cassava starch

Starches and gums are hydrocolloids frequently used in food systems to provide proper texture, moisture, and water mobility. Starch-gum interaction in food systems can change the starch granule swelling and its gelatinization and rheological properties. In this study, the effect of the addition of xanthan gum (XG), sodium carboxymethyl cellulose (SCMC), and carrageenan (CAR) at the concentrations of the 0.15, 0.25, 0.35 and 0.45% (w/v) on the pasting, thermal, and rheological properties of cassava starch was studied. The swelling power (SP) and the scanning electron microscopy (SEM) of the starch gels were also evaluated. The results obtained showed that xanthan gum (XG) had a strong interaction with the cassava starch penetrating between starch granules causing increase in pasting viscosities, SP, storage and loss (G', and G", respectively) modulus and reduction in the setback of the starch; sodium carboxymethyl cellulose (SCMC) greatly increased the pasting viscosities, the SP, and the storage and loss (G', and G", respectively) modulus of the starch-mixtures, mainly due to its greater capacity to hold water and not due to the interaction with cassava starch. Carrageenan (CAR) did not change any of the starch properties since there was no interaction between this gum and cassava starch at the concentrations used.